cutting out EGFP gene from pEGFP-N1 vector - what does dam- mean? (Jul/17/2008 )
we have a pEGFP-N1 vector with a gene of interest cloned into the MCS via EcoRI and BamH1; we wish to eliminate the EGFP but still using the CMV promoter and the gene of interest of the same vector; the 3´end of the EGFP gene is to digest with Xba I but it is methylated. CLONTECH recommends the following: "If you wish to digest the vector with this enzyme, you will need to transform the vector in to a dam- and make fresh DNA".
What is dam-?
Has anyone ever cut out EGFP from pEGP-N1, using which restriction enzymes?
most E coli strains contain the dam methylase (methylates G in the sequence GATC) and as a result, XbaI can no longer digest that site. We normally use the INV110 cells from Invitrogen to prepare non-methylated DNA, these are dam- and dcm-